Diabetic retinopathy, the leading cause of new blindness in individuals between the ages of 20-74 (1), is a disease process for which the pathological (anatomic) stages are well described. The underlying mechanisms, however, remain ill-defined or unknown. The plasminogen activator system consists of the proteases, tissue type-plasminogen activator and urokinase, and their natural inhibitors, plasminogen activator InhIbitor-1 (PAI-1) and PAI-2. The plasminogen activator system controls fibrinolysis and regulates extracellular matrix proteolysis. Disturbances in this system, in particular the overexpression of PAI by endothelial cells, in response to peptide hormones, growth factors, and cytokines, may be relevant to the pathogenesis of diabetic retinopathy. Aim 1. involves testing the hypothesis that elevated PAI levels result in the basement membrane thickening seen in diabetic retinal vessels. This will be tested by: A. Determining whether PAIs are expressed in greater quantities in the retinal capillaries of diabetics utilizing immunocytochemical studies. B. Assessing whether the exogenous administration of a synthetic PAI results in basement membrane thickening in the retinal vasculature. We will use direct intravitreal administration of synthetic PAIs followed by morphological studies. C. Determining whether overexpression of PAI-1 in the retinal vasculature results in similar retinal pathology to that observed in the diabetic. We will use morphometric analysis of capillaries in ocular tissue from transgenic mice overexpressing PAI-1 Aim 2. involves testing the hypothesis that the absence of PAI-1 makes the retinal vasculature resistant to developing diabetes induced changes. Utilizing PAI-1 deficient mice made diabetic by administration of streptozotocin. Aim 3. involves testing the hypothesis that IGF-l, regulated by the IGF binding proteins, and selected cytokines act as modulators of PAI-1 expression and that the diabetic state affects IGF-l regulation within the retina. These studies will utilize human retinal endothelial cells and donor human ocular tissue. Through the studies proposed in this application an understanding of the molecular mechanisms of PAI regulation by the retinal vasculature will be gained. The studies are critical to determining appropriate therapeutic interventions, including the use of agents such as growth factor inhibitors, anti-inflammatory agents, and agents such as metformin, that decrease PAI levels, in the management of diabetic patients before the development of clinically apparent background retinopathy.